Diversity matters: dynamic simulation of distributed bacterial states in suspended growth biological wastewater treatment systems

A MATLAB-based program was developed to simulate the distributions of states and behaviors of microbial storage product-accumulating bacteria in suspended growth systems. Currently available computer simulators of these systems predict dynamic behaviors by numerically solving differential biokinetic equations using average, or "lumped" system states (e.g., average microbial storage products concentrations). However, individual bacterial states are expected to diverge from average values, in part because individuals can have different hydrodynamic histories in terms of their residence times in upstream completely mixed flow reactors. The distributed state simulation program presented in this paper (DisSimulator 1.0) tracks individual bacteria as they move through a completely mixed reactor system. The program was evaluated for competition between polyphosphate-accumulating organisms (PAOs) and non-polyphosphate-accumulating heterotrophs in an enhanced biological phosphorus removal (EBPR) system for treatment of domestic wastewater. For identical systems and process conditions, simulations accounting for distributed states predicted larger anaerobic and aerobic solids residence time requirements for successful EBPR than did simulations using the lumped approach. One reason for this was that distributed simulations predicted large numbers of the PAOs were essentially inactive due to depleted or maximized storage product contents, while the lumped simulations predicted homogenous, 100% active PAO populations. Characteristic state profile shapes developed rapidly and were stable as total population numbers changed. Lumped state assumptions were demonstrated to produce large errors in predictions of EBPR system performance, and so consideration of distributed states may improve the accuracy of microbial storage products-based process simulations in systems with completely mixed hydrodynamics.     

Process hydraulics, distributed bacterial states, and biological phosphorus removal from wastewater

Hydraulic characteristics of biological wastewater treatment systems were shown to affect bacterial state distributions and system performance through mathematical simulations. The term "state" is used here to mean the microbial storage product and biomass content of a bacterium. The traditional approach to simulating biological treatment processes assumes "lumped" (average) states, rather than accounting for variable states across bacterial populations. Distributed states were previously suggested as critical to enhanced biological phosphorus removal (EBPR), but the factors that cause distributed states were not evaluated. A primary driver for distributed state development is variable hydraulic experiences of bacteria as they cycle through completely mixed reactors, and so process characteristics that affect hydraulics were hypothesized to affect state distributions. Two design characteristics affecting system hydraulics were evaluated using a new distributed state simulation program (DisSimulator 1.0): total hydraulic residence time (HRT) and numbers of reactors in series. Distributed predictions consistently predicted worse EBPR performance than did the lumped approach. Increasing HRTs (with constant solids retention times) tended to increase state distributions, to increase the differences between lumped and distributed simulation predictions, and to decrease predicted EBPR performance. As the numbers of reactors in series increased, distributed predictions tended to converge with lumped simulation predictions. Distributed simulations tended to predict a greater benefit to using reactors in series than did lumped simulations. This work provides guidance for new strategies to improve EBPR by minimizing state distributions. The targeted hydraulic characteristics may be more important to EBPR than previously recognized due to their effects on distributed states.     

A review and update of the microbiology of enhanced biological phosphorus removal in wastewater treatment plants

Enhanced biological phosphorus removal (EBPR) from wastewater can be more-or-less practically achieved but the microbiological and biochemical components are not completely understood. EBPR involves cycling microbial biomass and influent wastewater through anaerobic and aerobic zones to achieve a selection of microorganisms with high capacity to accumulate polyphosphate intracellularly in the aerobic period. Biochemical or metabolic modelling of the process has been used to explain the types of carbon and phosphorus transformations in sludge biomass. There are essentially two broad-groupings of microorganisms involved in EBPR. They are polyphosphate accumulating organisms (PAOs) and their supposed carbon-competitors called glycogen accumulating organisms (GAOs). The morphological appearance of microorganisms in EBPR sludges has attracted attention. For example, GAOs as tetrad-arranged cocci and clusters of coccobacillus-shaped PAOs have been much commented upon and the use of simple cellular staining methods has contributed to EBPR knowledge. Acinetobacter and other bacteria were regularly isolated in pure culture from EBPR sludges and were initially thought to be PAOs. However, when contemporary molecular microbial ecology methods in concert with detailed process performance data and simple intracellular polymer staining methods were used, a betaproteobacteria called ‘Candidatus Accumulibacter phosphatis’ was confirmed as a PAO and organisms from a novel gammaproteobacteria lineage were GAOs. To preclude making the mistakes of previous researchers, it is recommended that the sludge ‘biography’ be well understood – i.e. details of phenotype (process performance and biochemistry) and microbial community structure should be linked. This revised version was published online in August 2006 with corrections to the Cover Date.     

Competition between polyphosphate and glycogen accumulating organisms in enhanced biological phosphorus removal systems with acetate and propionate as carbon sources

Enhanced biological phosphorus removal (EBPR) is a widely used process for achieving phosphorus removal from wastewater. A potential reason for EBPR failure is the undesirable growth of glycogen accumulating organisms (GAOs), which can compete for carbon sources with the bacterial group responsible for phosphorus removal from wastewater: the polyphosphate accumulating organisms (PAOs). This study investigates the impact of carbon source on EBPR performance and the competition between PAOs and GAOs. Two sequencing batch reactors (SBRs) were operated during a 4-6 month period and fed with a media containing acetate or propionate, respectively, as the sole carbon source. It was found that the acetate fed SBR rarely achieved a high level of phosphorus removal, and that a large portion of the microbial community was comprised of "Candidatus Competibacter phosphatis", a known GAO. The propionate fed SBR, however, achieved stable phosphorus removal throughout the study, apart from one brief disturbance. The bacterial community of the propionate fed SBR was dominated by "Candidatus Accumulibacter phosphatis", a known PAO, and did not contain Competibacter. In a separate experiment, another SBR was seeded with a mixture of PAOs and a group of alphaproteobacterial GAOs, both enriched with propionate as the sole carbon source. Stable EBPR was achieved and the PAO population increased while the GAOs appeared to be out-competed. The results of this paper suggest that propionate may provide PAOs with a selective advantage over GAOs in the PAO-GAO competition, particularly through the minimisation of Competibacter. Propionate may be a more suitable substrate than acetate for enhancing phosphorus removal in EBPR systems.     

强化生物除磷系统主要微生物及其代谢机理研究进展

强化生物除磷(enhanced biological phosphorus removal,EBPR)工艺在废水除磷处理中应用广泛.主要功能微生物及其代谢机理的研究是有效调控EBPR工艺稳定运行与效能提升的基础.本文选取EBPR系统中最主要的两类微生物(聚磷菌和聚糖菌),从底物吸收机制、糖酵解途径、TCA途径的贡献以及聚磷菌和聚糖菌的代谢相似性等方面对这些微生物的代谢机理进行综述,评价了分子生物学技术在研究EBPR系统微生物学及其代谢机理方面的应用现状,在此基础上对EBPR系统今后的研究方向进行了展望.
     

Microbial population analysis of nutrient removal-related organisms in membrane bioreactors

Membrane bioreactors (MBR) are an important and increasingly implemented wastewater treatment technology, which are operated at low food to microorganism ratios (F/M) and retain slow-growing organisms. Enhanced biological phosphorus removal (EBPR)-related organisms grow slower than ordinary heterotrophs, but have never been studied in detail in MBRs. This study presents a comprehensive analysis of the microorganisms involved in EBPR in pilot- and full-scale MBRs, using fluorescence in situ hybridization (FISH), as well as an overall assessment of other relevant microbial groups. The results showed that polyphosphate accumulating organisms (PAOs) were present at similar levels in all studied MBRs (10% ± 6%), even those without a defined anaerobic zone. Glycogen accumulating organisms were also detected, although rarely. The FISH results correlated well with the observed P removal performance of each plant. The results from this study suggest that a defined anaerobic zone is not necessarily required for putative PAO growth in MBRs, since polyphosphate storage may provide a selective advantage in fulfilling cell maintenance requirements in substrate-limited conditions (low F/M).     

The microbiology of biological phosphorus removal in activated sludge systems

Activated sludge systems are designed and operated globally to remove phosphorus microbiologically, a process called enhanced biological phosphorus removal (EBPR). Yet little is still known about the ecology of EBPR processes, the microbes involved, their functions there and the possible reasons why they often perform unreliably. The application of rRNA-based methods to analyze EBPR community structure has changed dramatically our understanding of the microbial populations responsible for EBPR, but many substantial gaps in our knowledge of the population dynamics of EBPR and its underlying mechanisms remain. This review critically examines what we once thought we knew about the microbial ecology of EBPR, what we think we now know, and what still needs to be elucidated before these processes can be operated and controlled more reliably than is currently possible. It looks at the history of EBPR, the currently available biochemical models, the structure of the microbial communities found in EBPR systems, possible identities of the bacteria responsible, and the evidence why these systems might operate suboptimally. The review stresses the need to extend what have been predominantly laboratory-based studies to full-scale operating plants. It aims to encourage microbiologists and process engineers to collaborate more closely and to bring an interdisciplinary approach to bear on this complex ecosystem.     

Influence of sludge retention time on tolerance of copper toxicity for polyphosphate accumulating organisms linked to polyhydroxyalkanoates metabolism and phosphate removal

This study explored the influence of sludge retention time (SRT) on tolerance of copper invasion for polyphosphate accumulating organisms (PAOs) in an enhanced biological phosphorus removal (EBPR). The experimental data showed the anaerobic polyhydroxyalkanoates (PHA) storage for the sludge at 10d SRT was less influenced by copper invasion than those at 5d and 15d SRTs. The reaction of PAOs aerobically taking up phosphate for the sludge at 5d or 15d SRT almost ceased at 2 mg Cu L⁻¹, whereas PAOs in the sludge at 10d SRT retained half of the ability to take up phosphate. Both the PHAs degradation and synthesis rates decreased with increasing copper concentration, regardless of the SRTs. However, the copper inhibition of the former was greater than that of the later.     

Microbial selection of polyphosphate-accumulating bacteria in activated sludge wastewater treatment processes for enhanced biological phosphate removal

Activated sludge processes with alternating anaerobic and aerobic conditions (the anaerobic-aerobic process) have been successfully used for enhanced biological phosphate removal (EBPR) from wastewater. It is known that polyphosphate-accumulating bacteria (PAB) play an essential role for EBPR in the anaerobic-aerobic process. The present paper reviews limited information available on the metabolism and the microbial community structure of EBPR, highlighting the microbial ecological selection of PAB in EBPR processes. Exposure of microorganisms to alternate carbon-rich anaerobic environments and carbon-poor aerobic environments in the anaerobic-aerobic process induces the key metabolic characteristics of PAB, which include organic substrate uptake followed by its conversion to stored polyhydroxyalkanoate (PHA) and hydrolysis of intracellular polyphosphate accompanied by subsequent Pi release under anaerobic conditions. Intracellular glycogen is assumed to function as a regulator of the redox balance in the cell. Storage of glycogen is a key strategy for PAB to maintain the redox balance in the anaerobic uptake of various organic substrates, and hence to win in the microbial selection. Acinetobacter spp., Microlunatus phosphovorus, Lampropedia spp., and the Rhodocyclus group have been reported as candidates of PAB. PAB may not be composed of a few limited genospecies, but involve phylogenetically and taxonomically diverse groups of bacteria. To define microbial community structure of EBPR processes, it is needed to look more closely into the occurrence and behavior of each species of PAB in various EBPR processes mainly by molecular methods because many of PAB seem to be impossible to culture.     

Characterizing the biochemical activity of full-scale enhanced biological phosphorus removal systems: A comparison with metabolic models

The metabolism of polyphosphate accumulating organisms (PAOs) has been widely studied through the use of lab-scale enrichments. Various metabolic models have been formulated, based on the results from lab-scale experiments using enriched PAO cultures. A comparison between the anaerobic stoichiometry predicted by metabolic models with that exhibited by full-scale sludge in enhanced biological phosphorus removal (EBPR) wastewater treatment plants (WWTPs) was performed in this study. Batch experiments were carried out with either acetate or propionate as the sole carbon source, using sludges from two different EBPR-WWTPs in Australia that achieved different phosphorus removal performances. The results support the hypothesis that the anaerobic degradation of glycogen is the primary source of reducing equivalents generated by PAOs, however, they also suggested a partial contribution of the tricarboxylic acid (TCA) cycle in some cases. The experimental results obtained when acetate was the carbon source suggest the involvement of the modified succinate-propionate pathway for the generation of poly-beta-hydroxyvalerate (PHV). Overall, the batch test results obtained from full-scale EBPR sludge with both substrates were generally well described by metabolic model predictions for PAOs.     

Monitoring the impact of bioaugmentation on the start up of biological phosphorus removal in a laboratory scale activated sludge ecosystem

The acclimatisation of activated sludge to enhanced biological phosphorus removal (EBPR) conditions requires a period of about 40–100 days but its output remains hazardous. The impact of bioaugmentation on the start-up of a laboratory scale EBPR sequencing batch reactor was evaluated by process parameters measurement and microbial community dynamics monitoring using 16S rDNA targeted polymerase chain reaction-single strand conformation polymorphism electrophoresis (PCR-SSCP). Bioaugmentation: (1) speeded up the installation of good and stable EBPR in the bioaugmented reactor by about 15 days; (2) correlated with the transient enrichment of the sludge in the added microbial populations; and (3) favoured the long-term enrichment of the sludge in the phosphorus-accumulating organism (PAO) Candidatus Accumulibacter phosphatis. However, despite a lag time period, the control non-bioaugmented reactor ended up with comparable reactor parameters and microbial community evolution, suggesting that the same PAO populations were already present from the beginning in the original non-P-accumulating seed sludge. The potential of a true installation of the added microbial populations within the bioaugmented reactor compared to their substitution by indigenous similar populations is discussed. Competition between PAOs and the antagonistic glycogen accumulating organism Candidatus Competibacter phosphatis is also highlighted during EBPR start-up.     

Aerobic phosphorus release linked to acetate uptake in bio-P sludge: process modeling using oxygen uptake rate

The main processes involved in enhanced biological phosphorus removal (EBPR) under anaerobic and subsequently aerobic conditions are widely described in the literature. Polyphosphate accumulating organisms (PAO) are the organisms responsible for this process. However, the mechanisms of PAO are not fully established yet under conditions that differ from the classical anaerobic/aerobic conditions. In this work, we made a comparison between the behavior of PAO under classical EBPR conditions and its behavior when consuming substrate under only aerobic conditions. In addition, oxygen uptake rate (OUR) was measured in the set of experiments under aerobic conditions to improve the characterization of the process. A kinetic and stoichiometric model based on Activated Sludge Model No.2 (ASM2) and including glycogen economy (AnOx model), calibrated for classical anaerobic/aerobic conditions, was not able to describe the experimental data since it underestimated the acetate consumption, the PHB storage, and the OUR. Two different hypotheses for describing the experimental measurements were proposed and modeled. Both hypotheses considered that PAO, under aerobic conditions, uptake acetate coupled to PHB storage, glycogen degradation, and phosphorus release as in anaerobic conditions. Moreover, the first hypothesis (PAO-hypothesis) considered that PAO were able to store acetate as PHB linked to oxygen consumption and the second one (OHO hypothesis) considered that this storage was due to ordinary heterotrophic organisms (OHO). Both hypotheses were evaluated by simulation extending the AnOx model with additional equations. The main differences observed were the predictions for PHB degradation during the famine phase and the OUR profile during both feast and famine phases. The OHO hypothesis described the experimental profiles more accurately than the PAO hypothesis.     

Post-anoxic denitrification driven by PHA and glycogen within enhanced biological phosphorus removal

The objective of this research was to interrogate and develop a better understanding for a process to achieve post-anoxic denitrification without exogenous carbon augmentation within enhanced biological phosphorus removal (EBPR). Sequencing batch reactors fed real wastewater and seeded with mixed microbial consortia were operated under variable anaerobic-aerobic-anoxic and organic carbon loading conditions. The process consistently achieved phosphorus and nitrogen removal, while the observed specific denitrification rates were markedly higher than expected for post-anoxic systems operated without exogenous organic carbon addition. Investigations revealed that post-anoxic denitrification was predominantly driven by glycogen, an intracellular carbon storage polymer associated with EBPR; moreover, glycogen reserves can be significantly depleted post-anoxically without compromising EBPR. Success of the proposed process is predicated on providing sufficient organic acids in the influent wastewater, such that residual nitrate carried over from the post-anoxic period is reduced and polyhydroxyalkanoate (PHA) synthesis occurs.     

Radiolabelled proteomics to determine differential functioning of Accumulibacter during the anaerobic and aerobic phases of a bioreactor operating for enhanced biological phosphorus removal

Proteins synthesized by the mixed microbial community of two sequencing batch reactors run for enhanced biological phosphorus removal (EBPR) during aerobic and anaerobic reactor phases were compared, using mass spectrometry‐based proteomics and radiolabelling. Both sludges were dominated by polyphosphate‐accumulating organisms belonging to Candidatis Accumulibacter and the majority of proteins identified matched closest to these bacteria. Enzymes from the Embden–Meyerhof–Parnas pathway were identified, suggesting this is the major glycolytic pathway for these Accumulibacter populations. Enhanced aerobic synthesis of glyoxylate cycle enzymes suggests this cycle is important during the aerobic phase of EBPR. In one sludge, several TCA cycle enzymes showed enhanced aerobic synthesis, suggesting this cycle is unimportant anaerobically. The second sludge showed enhanced synthesis of TCA cycle enzymes under anaerobic conditions, suggesting full or partial TCA cycle operation anaerobically. A phylogenetic analysis of Accumulibacter polyphosphate kinase genes from each sludge demonstrated different Accumulibacter populations dominated the two sludges. Thus, TCA cycle activity differences may be due to Accumulibacter strain differences. The major fatty acids present in Accumulibacter‐dominated sludge include palmitic, hexadecenoic and cis‐vaccenic acid and fatty acid content increased by approximately 20% during the anaerobic phase. We hypothesize that this is associated with increased anaerobic phospholipid membrane biosynthesis, to accommodate intracellular polyhydroxyalkanoate granules.     

Polyhydroxyalkanoate synthesis using different carbon sources by two enhanced biological phosphorus removal microbial communities

Polyhydroxyalkanoate (PHA) is a biodegradable plastic synthesised by bacteria as energy and carbon storage material. PHA production is mostly based on pure cultures operated under sterile conditions, which increase the costs of this biopolymer. The use of inexpensive mixed culture biomass, such as activated sludge, to produce biodegradable plastics from renewable waste streams has been proposed as an alternative.The effect of carbon sources (acetate, propionate, butyrate and glucose) on the type and quantity of PHA synthesis obtained with different enhanced biological phosphorus removal (EBPR) microbial communities enriched with acetate and propionate are reported in this work. Two sequencing batch reactors (SBRs) were seeded with biomass withdrawn from a non-EBPR wastewater treatment plant (WWTP). The same operational conditions were kept, but using acetate or propionate as the sole carbon source of each reactor. These conditions produced two microbial communities with different P-removal capacity. The results presented in this study show the effect of the carbon source on the PHA composition (amount of polyhydroxybutyrate (PHB), polyhydroxyvalerate (PHV) and polyhydroxy-2-methylvalerate (PH2MV)), which differed not just between substrates but also between the two EBPR communities used. In addition, some monomers not always analysed contribute significantly to the total amount of PHA, especially when using butyrate, showing that if they are not considered this can lead to erroneous calculated yields.     

Effect of nitrite from nitritation on biological phosphorus removal in a sequencing batch reactor treating domestic wastewater

Although nitrite effect on enhanced biological phosphorus removal (EBPR) has been previously studied, very limited research has been undertaken about the effect of nitrite accumulation caused by nitritation on EBPR. This paper focused on nitrite effect from nitritation on EBPR in a sequencing batch reactor treating domestic wastewater. Results showed that nitrite of below 10 mg/L did not inhibit P-uptake and release; whereas EBPR deterioration was observed when nitrite accumulation reached 20 mg/L. Due to P-uptake prior to nitritation, nitrite of 20 mg/L has no effect on aerobic P-uptake. The main reason leading to EBPR deterioration was the competition of carbon source. Batch tests were conducted to investigate nitrite effect on anaerobic P-release. Under sufficient carbon source, nitrite of 30 mg/L had no impact on poly-β-hydroxyalkanoate (PHA) storage; contrarily, under insufficient carbon source, denitrifiers competing for carbon source with phosphorus accumulating organisms resulted in decrease of PHA synthesis and P-release.     

Planning of distributed data production for High Energy and Nuclear Physics

Modern experiments in High Energy and Nuclear Physics heavily rely on distributed computations using multiple computational facilities across the world. One of the essential types of the computations is a distributed data production where petabytes of raw files from a single source has to be processed once (per production campaign) using thousands of CPUs at distant locations and the output has to be transferred back to that source. The data distribution over a large system does not necessary match the distribution of storage, network and CPU capacity. Therefore, bottlenecks may appear and lead to increased latency and degraded performance. In this paper we propose a new scheduling approach for distributed data production which is based on the network flow maximization model. In our approach a central planner defines how much input and output data should be transferred over each network link in order to maximize the computational throughput. Such plans are created periodically for a fixed planning time interval using up-to-date information on network, storage and CPU resources. The centrally created plans are executed in a distributed manner by dedicated services running at participating sites. Our simulations based on the log records from the data production framework of the experiment STAR (Solenoid Tracker at RHIC) have shown that the proposed model systematically provides a better performance compared to the simulated traditional techniques.     

聚磷菌和聚糖菌的竞争影响因素研究进展

目前, 强化生物除磷工艺(EBPR)以其经济有效而得到广泛的应用, 该工艺关键在于聚磷菌的富集。然而已经发现, 有一类细菌—聚糖菌(GAOs)能够和聚磷菌(PAOs)竞争, 导致除磷效果恶化。关于PAOs-GAOs的竞争, 研究已经很多, 但是其结论有趋同也有矛盾, 有必要对此进行分析讨论。根据近年来国内外的相关报道, 阐述了聚磷菌与聚糖菌的竞争影响因素, 其中碳磷比、碳源种类、温度、pH值是关键因素, 而污泥龄、溶解氧以及水力停留时间等因素对于PAOs和GAOs的竞争也起一定的作用。此外, 在EBPR系统中, 缺氧条件下, 存在反硝化聚磷菌(DPB)和反硝化聚糖菌(DGAO)也会对聚磷菌富集和系统除磷产生影响。最后对EBPR系统未来的发展方向进行了展望。